title: A genetic interaction between NDPK and AMPK in Dictyostelium discoideum that affects motility, growth and development.
abstract: Many of the expanding roles of nucleoside diphosphate kinase have been attributed to its ability to interact with other proteins. One proposal is an interaction with the cellular energy sensor AMP-activated protein kinase, and here, we apply the simple eukaryotic organism, Dictyostelium discoideum as a test model. Stable cotransformants were created in which NDPK expression was knocked down by antisense inhibition, and AMPK activity was chronically elevated either by constitutive overexpression of its active, catalytic domain (AMPKαT) or as a result of mitochondrial dysfunction (created by antisense inhibition of expression of a mitochondrial chaperone protein, chaperonin 60). To investigate a biochemical interaction, transformants were created which contained constructs expressing FLAG-NDPK and hexahistidine-tagged full-length AMPK or AMPKαT. The protein extract from these transformants was used in coimmunoprecipitations. Knock down of NDPK expression suppressed the phenotypic defects that are caused by AMPK hyperactivity resulting either from overexpression of AMPKαT or from mitochondrial dysfunction. These included rescue of defects in slug phototaxis, fruiting body morphology and growth in a liquid medium. Coimmunoprecipitation experiments failed to demonstrate a biochemical interaction between the two proteins. The results demonstrate a genetic interaction between NDPK and AMPK in Dictyostelium in that NDPK is required for the phenotypic effects of activated AMPK. Coimmunoprecipitations suggest that this interaction is not mediated by a direct interaction between the two proteins.
authors.affiliation: Department of Microbology, La Trobe University, Melbourne, VIC, 3086, Australia. S.Annesley@latrobe.edu.au
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Annesley
authors.forename: Ruzica
authors.initials: R
authors.lastname: Bago
authors.forename: Anil
authors.initials: A
authors.lastname: Mehta
authors.forename: Paul R
authors.initials: PR
authors.lastname: Fisher
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics*; D004023:Dictyostelium / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D015971:Gene Expression Regulation, Enzymologic; D008954:Models, Biological; D009068:Movement; D009701:Nucleoside-Diphosphate Kinase / Q000235:genetics*
chemical_list: D055372:AMP-Activated Protein Kinases; D009701:Nucleoside-Diphosphate Kinase